Discovery of Enzyme Structure Linked to Cancer May Lead to New Therapies

Scientists at the University of California, Riverside, say they have discovered the structure of a novel RNA-modifying enzyme, ZCCHC4, and identified the mechanism that controls how this enzyme recognizes its substrate. ZCCHC4 influences cell proliferation and has been linked to cancers. It uniquely introduces one kind of RNA modification, N6-methyladenosine (m6A), into ribosomes, according to the team.

The group’s study “Structure and regulation of ZCCHC4 in m6A-methylation of 28S rRNA”, published in Nature Communications, explains how protein machineries in cells are regulated to target RNA molecules for m6A modification.

Jikui Song, PhD, an associate professor of biochemistry, who led the study, explained ZCCHC4 controls protein synthesis and cell proliferation by introducing an m6A modification into ribosomes. ZCCHC4, he added, is overexpressed in tumors associated with hepatocellular carcinoma, the most common type of primary liver cancer.

“This is the first time anyone has determined the crystal structure of ZCCHC4,” Song said. “Our discovery can be used for structure-based drug design against cancers and lead to a better understanding of how m6A, a modification associated with numerous biological processes, is installed on ribosomal RNA.”

The m6A modification has received enormous attention in recent years due to the important role it plays in RNA metabolism and biology. How this modification is dynamically programmed and distributed in cells, however, remains poorly understood.

“The structure of ZCCHC4 provides an understanding of how this enzyme is wired to specifically act on ’28S ribosomal RNA,'” Song continued, noting a ribosome is assembled with differently sized subunits. 28S ribosomal RNA refers to the RNA component in the 28S ribosomal subunit. “We now understand that this enzyme is controlled by an ‘autoinhibitory’ mechanism that has been observed in many other cellular processes.”

To crack the structure of ZCCHC4, Song’s team first produced an enzymatically active and structurally rigid ZCCHC4 fragment. The researchers then coaxed this protein to crystallize. Finally, they diffracted the crystals using x-rays and analyzed the data, which led to the eventual discovery of ZCCHC4’s structure.

“N6-methyladenosine (m6A) modification provides an important epitranscriptomic mechanism that critically regulates RNA metabolism and function. However, how m6A writers attain substrate specificities remains unclear. We report the 3.1 Å-resolution crystal structure of human CCHC zinc finger-containing protein ZCCHC4, a 28S rRNA-specific m6A methyltransferase, bound to S-adenosyl-L-homocysteine. The methyltransferase (MTase) domain of ZCCHC4 is packed against N-terminal GRF-type and C2H2 zinc finger domains and a C-terminal CCHC domain, creating an integrated RNA-binding surface. Strikingly, the MTase domain adopts an autoinhibitory conformation, with a self-occluded catalytic site and a fully-closed cofactor pocket,” the investigators wrote.

“Mutational and enzymatic analyses further substantiate the molecular basis for ZCCHC4-RNA recognition and a role of the stem-loop structure within the substrate in governing the substrate specificity. Overall, this study unveils unique structural and enzymatic characteristics of ZCCHC4, distinctive from what was seen with the METTL family of m6A writers, providing the mechanistic basis for ZCCHC4 modulation of m6A RNA methylation.”

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